The dopamine- and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32) is critical to the pathogenesis of drug addiction by modulating both transcriptional and post-translational events in dopaminergic neurons of the brain. DARPP-32 is a central mediator of the activity of the extracellular signal-regulated kinase (ERK) signaling cascade, the activation of which results in a stimulus-reward mechanism that propagates the cycle of addictive behaviors. We hypothesize that inhibition of the gene expression of DARPP-32 by dopaminergic neurons in vivo using specific short interfering RNA (siRNA) will abrogate the behaviors associated with drug addiction. Silencing of DARPP-32 using an innovative, nanoparticle-based delivery system employing gold nanorods (GNR) complexed with siRNA specific for DARPP-32 plus tranferrin to specifically cross the blood-brain barrier (BBB) and target neurons may provide an innovative gene therapy for drug addiction. In this proposal, we seek to chemically characterize the GNR and develop stable nanoplexes containing siRNA and transferrin, which can effectively silence DARPP-32 gene expression in vivo in the brains of an animal model of opiate addiction, thereby suppressing addictive behavior. Additionally, we will generate mechanism-based pharmacodynamics (PD) models for evaluating DARPP-32 signaling and its implications in drug dynamics. Based on the above hypothesis, the following specific aims are proposed. Aim 1: To optimize the conditions for the efficient delivery of gold nanoplexes across the BBB, in vivo, in an established rat model of chronic opiate abuse using a GNR-DARPP- 32siRNA nanoplex and to evaluate its effect on both the gene silencing and the protein expression of DARPP-32, as well as of other key signal transduction molecules such as CREB and PP-1 that are part of the dopaminergic signaling pathway and control transcriptional events down stream of DARPP-32. Aim 2: To investigate the effects of delivery of GNR-DARPP-32siRNA nanoplexes in vivo on opiate withdrawal behavior in an established rat model of chronic opiate abuse. Aim 3: To develop predictive models of DARPP-32 signaling using a novel multi-scale systems analysis approach to determine the complex interrelationships between cell systems and drug factors that may influence drug addiction and its treatment. These nanoplexes have the potential to not only be a powerful new tool to study and manipulate, at the molecular level, various signal transduction pathways in the human brain that affect addictive behavior, but also facilitate the development of GNR conjugated siRNA-based therapy for drug addiction. Further the pharmacodynamics models that will emerge from this study will help describe, understand, and predict the complex and dynamic interactions between neurotransmitters and their receptors and complex mechanisms of action that underlie the process of drug addiction. PUBLIC HEALTH RELEVANCE: Drugs of abuse act on the dopaminergic system of the brain and perturb the function of dopamine and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32), which is critical to the pathogenesis of drug addiction. The current project involves silencing of DARPP-32 gene expression using innovative nanotechnology based siRNA therapeutics delivered to the brain in vivo as a new approach for the treatment of drug addiction. Additionally, we will generate pharmacodynamics (PD) models for evaluating DARPP-32 signaling and its implications in drug addiction dynamics.